Monoclonal antibodies are essential reagents for deciphering gene or protein function and have been a fruitful source of therapeutic and diagnostic agents. However, the use of anticarbohydrate antibodies to target glycans for these purposes has been less successful. Glycans contain less hydrophobic functionality than do proteins or nucleic acids, thus individual glycan-antibody interactions are relatively weak. Information encoded by glycans often involves subtle variations of branched oligosaccharides that cannot be detected with conventional antibodies. It has therefore been difficult to obtain reagents that enable a complete understanding of biological glycan function. Changes in cell surface glycan composition are associated with cancer (and other disease states); therefore, general methods to identify specific and high- affinity glycan antibodies would greatly facilitate cancer research and provide new avenues for cancer therapies and diagnostics. We propose to develop methods to identify such reagents using novel antibody phage display technologies. It has recently become possible to select specific and high affinity antibodies for virtually any protein antigen from phage libraries that bear tailored diversity elements encoded by synthetic DNA ('synthetic antibodies'). This innovative technology platform obviates the requirement for animal immunization, thereby circumventing many limitations of traditional hybridoma methods. We will use the unique architectural scaffold of 2G12, an antibody that targets oligomannoses on the HIV-1 glycoprotein gp120, as the template for our design. The two heavy chain variable domains of 2G12 IgG exchange positions to create an extended recognition surface containing four oligomannose binding sites per IgG molecule. We have developed a phage vector that allows the functional display of the 2G12 scaffold. We will prepare synthetic 2G12 antibody libraries to determine minimal physicochemical requirements for high affinity glycan recognition in this scaffold. Next, we will use the information gained from these studies to identify novel 2G12 variants with altered specificity profiles for tumor-associated glycan targets. This innovative strategy will result in novel cancer antibodies that accelerate cancer research and pave the way for development of new cancer therapies and diagnostics.